Yonglan Zheng

Whole-exome and targeted gene sequencing of gallbladder carcinoma identifies recurrent mutations in the ErbB pathway

Individuals with gallbladder carcinoma (GBC), the most aggressive malignancy of the biliary tract, have a poor prognosis. Here we report the identification of somatic mutations for GBC in 57 tumor-normal pairs through a combination of exome sequencing and ultra-deep sequencing of cancer-related genes. The mutation pattern is defined by a dominant prevalence of C>T mutations at TCN sites. Genes with a significant frequency (false discovery rate (FDR) < 0.05) of non-silent mutations include TP53 (47.1%), KRAS (7.8%) and ERBB3 (11.8%). Moreover, ErbB signaling (including EGFR, ERBB2, ERBB3, ERBB4 and their downstream genes) is the most extensively mutated pathway, affecting 36.8% (21/57) of the GBC samples. Multivariate analyses further show that cases with ErbB pathway mutations have a worse outcome (P = 0.001). These findings provide insight into the somatic mutational landscape in GBC and highlight the key role of the ErbB signaling pathway in GBC pathogenesis.

Evaluation of 19 susceptibility loci of breast cancer in women of African ancestry

Multiple breast cancer susceptibility loci have been identified in genome-wide association studies (GWAS) in populations of European and Asian ancestry using array chips optimized for populations of European ancestry. It is important to examine whether these loci are associated with breast cancer risk in women of African ancestry. We evaluated 25 single nucleotide polymorphisms (SNPs) at 19 loci in a pooled case-control study of breast cancer, which included 1509 cases and 1383 controls. Cases and controls were enrolled in Nigeria, Barbados and the USA; all women were of African ancestry. We found significant associations for three SNPs, which were in the same direction and of similar magnitude as those reported in previous fine-mapping studies in women of African ancestry. The allelic odds ratios were 1.24 [95% confidence interval (CI): 1.04-1.47; P = 0.018] for the rs2981578-G allele (10q26/FGFR2), 1.34 (95% CI: 1.10-1.63; P = 0.0035) for the rs9397435-G allele (6q25) and 1.12 (95% CI: 1.00-1.25; P = 0.04) for the rs3104793-C allele (16q12). Although a significant association was observed for an additional index SNP (rs3817198), it was in the opposite direction to prior GWAS studies. In conclusion, this study highlights the complexity of applying current GWAS findings across racial/ethnic groups, as none of GWAS-identified index SNPs could be replicated in women of African ancestry. Further fine-mapping studies in women of African ancestry will be needed to reveal additional and causal variants for breast cancer.

Screening RAD51C nucleotide alterations in patients with a family history of breast and ovarian cancer.

It has been reported that one biallelic missense mutation in the RAD51C gene was found in a Fanconi anemia-like disorder and six monoallelic pathogenic mutations were identified in 480 BRCA1/2 negative breast and ovarian cancer pedigrees but not in 620 pedigrees with breast cancer only. Additionally, the RAD51C gene was reported to be involved in gene fusion events in the MCF-7 breast cancer cell line. We performed complete sequencing and fusion gene breakpoint screening to detect deleterious mutations and chromosomal structure change in the RAD51C gene. Ninety-two hereditary gynecological cancer patients with a family history of breast and ovarian cancer but not carrying BRCA1/2 mutations were studied. In addition, 46 breast cancer cell lines were screened for the gene fusion events. Ten DNA sequence variants but no deleterious mutations were identified. We did not observe the occurrence of the known gene fusion either. We were unable to confirm the contribution of the RAD51C gene to hereditary breast and ovarian cancer (HBOC) in this relatively small cohort. Nonetheless, larger studies in diverse populations to fully investigate the mutation spectrum of the RAD51C gene are needed.

High prevalence of BRCA1 and BRCA2 mutations in unselected Nigerian breast cancer patients

Inherited mutations in the BRCA1 and BRCA2 genes are the strongest genetic predictors of breast cancer and are the primary causes of familial breast/ovarian cancer syndrome. The frequency, spectrum and penetrance of mutant BRCA1/BRCA2 alleles have been determined for several populations, but little information is available for populations of African ancestry, who suffer a disproportionate burden of early onset breast cancer. We have performed complete sequence analysis of all BRCA1 and BRCA2 exons and intron–exon boundaries for 434 Nigerian breast cancer patients from the University College Hospital in Ibadan, Nigeria. In contrast to previous suggestions that BRCA1/BRCA2 mutation frequencies are low or undetectable in African American populations, we find that Nigerian breast cancer patients have an exceptionally high frequency of BRCA1 and BRCA2 mutations (7.1 and 3.9%, respectively). Sixteen different BRCA1 mutations were detected, seven of which have never been reported previously, while thirteen different BRCA2 mutations were seen, six of which were previously unreported. Thus, our data support enrichment for genetic risk factors in this relatively young cohort. To improve breast cancer outcomes, we suggest that family‐based models of risk assessment and genetic counseling coupled with interventions to reduce breast cancer risk should be broadly disseminated in Nigeria and other underserved and understudied populations.

Novel germline PALB2 truncating mutations in African American breast cancer patients.

It has been demonstrated that the partner and localizer of breast cancer 2 (PALB2) acts as a bridging molecule between the breast cancer 1 (BRCA1) and BRCA2 proteins and is responsible for facilitating BRCA2‐mediated DNA repair. Truncating mutations in the PALB2 gene reportedly are enriched in patients with Fanconi anemia and breast cancer in various populations.

Genome-wide association studies in women of African ancestry identified 3q26.21 as a novel susceptibility locus for oestrogen receptor negative breast cancer.

Multiple breast cancer loci have been identified in previous genome-wide association studies, but they were mainly conducted in populations of European ancestry. Women of African ancestry are more likely to have young-onset and oestrogen receptor (ER) negative breast cancer for reasons that are unknown and understudied. To identify genetic risk factors for breast cancer in women of African descent, we conducted a meta-analysis of two genome-wide association studies of breast cancer; one study consists of 1,657 cases and 2,029 controls genotyped with Illumina’s HumanOmni2.5 BeadChip and the other study included 3,016 cases and 2,745 controls genotyped using Illumina Human1M-Duo BeadChip. The top 18,376 single nucleotide polymorphisms (SNP) from the meta-analysis were replicated in the third study that consists of 1,984 African Americans cases and 2,939 controls. We found that SNP rs13074711, 26.5 Kb upstream of TNFSF10 at 3q26.21, was significantly associated with risk of oestrogen receptor (ER)-negative breast cancer (odds ratio [OR]=1.29, 95% CI: 1.18-1.40; P = 1.8 × 10 − 8). Functional annotations suggest that the TNFSF10 gene may be involved in breast cancer aetiology, but further functional experiments are needed. In addition, we confirmed SNP rs10069690 was the best indicator for ER-negative breast cancer at 5p15.33 (OR = 1.30; P = 2.4 × 10 − 10) and identified rs12998806 as the best indicator for ER-positive breast cancer at 2q35 (OR = 1.34; P = 2.2 × 10 − 8) for women of African ancestry. These findings demonstrated additional susceptibility alleles for breast cancer can be revealed in diverse populations and have important public health implications in building race/ethnicity-specific risk prediction model for breast cancer.

Fine-mapping of Breast Cancer Genome-wide Association Studies Loci in Women of African Ancestry Identifies Novel Susceptibility Markers

Numerous single nucleotide polymorphisms (SNPs) associated with breast cancer susceptibility have been identified by genome-wide association studies (GWAS). However, these SNPs were primarily discovered and validated in women of European and Asian ancestry. Because linkage disequilibrium is ancestry-dependent and heterogeneous among racial/ethnic populations, we evaluated common genetic variants at 22 GWAS-identified breast cancer susceptibility loci in a pooled sample of 1502 breast cancer cases and 1378 controls of African ancestry. None of the 22 GWAS index SNPs could be validated, challenging the direct generalizability of breast cancer risk variants identified in Caucasians or Asians to other populations. Novel breast cancer risk variants for women of African ancestry were identified in regions including 5p12 (odds ratio [OR] = 1.40, 95% confidence interval [CI] = 1.11-1.76; P = 0.004), 5q11.2 (OR = 1.22, 95% CI = 1.09-1.36; P = 0.00053) and 10p15.1 (OR = 1.22, 95% CI = 1.08-1.38; P = 0.0015). We also found positive association signals in three regions (6q25.1, 10q26.13 and 16q12.1-q12.2) previously confirmed by fine mapping in women of African ancestry. In addition, polygenic model indicated that eight best markers in this study, compared with 22 GWAS-identified SNPs, could better predict breast cancer risk in women of African ancestry (per-allele OR = 1.21, 95% CI = 1.16-1.27; P = 9.7 × 10(-16)). Our results demonstrate that fine mapping is a powerful approach to better characterize the breast cancer risk alleles in diverse populations. Future studies and new GWAS in women of African ancestry hold promise to discover additional variants for breast cancer susceptibility with clinical implications throughout the African diaspora.

TERT Polymorphism rs2736100-C Is Associated with EGFR Mutation–Positive Non–Small Cell Lung Cancer

Purpose: EGF receptor (EGFR) mutation–positive (EGFRmut+) non–small cell lung cancer (NSCLC) may be a unique orphan disease. Previous studies suggested that the telomerase reverse transcriptase (TERT) gene polymorphism is associated with demographic and clinical features strongly associated with EGFR mutations, for example, adenocarcinoma histology, never-smoking history, and female gender. We aim to test the association between TERT polymorphism and EGFRmut+ NSCLC. Experimental Design: We conducted a genetic association study in Chinese patients with NSCLC (n = 714) and healthy controls (n = 2,520), between the rs2736100 polymorphism and EGFRmut+ NSCLC. We further tested the association between the EGFR mutation status and mean leukocyte telomere length (LTL). The potential function of rs2736100 in lung epithelial cells was also explored. Results: The rs2736100-C allele was significantly associated with EGFRmut+ NSCLC [OR, 1.52; 95% confidence interval (CI), 1.28–1.80; P = 1.6 × 10−6] but not EGFRmut− NSCLC (OR = 1.07, 95% CI, 0.92–1.24, P = 0.4). While patients with NSCLC as a whole have significantly longer LTL than healthy controls (P ≤ 10−13), the EGFRmut+ patients have even longer LTL than EGFRmut− patients (P = 0.008). Meanwhile, rs2736100 was significantly associated with TERT mRNA expression in both normal and tumor lung tissues. All results remained significant after controlling for age, gender, smoking status, and histology (P < 0.05 for all tests). Moreover, the rs2736100 DNA sequence has an allele-specific affinity to nuclear proteins extracted from lung epithelial cells, which led to an altered enhancer activity of the sequence in vitro. Conclusions: Our study suggests that telomerase and telomere function may be essential for carcinogenesis of EGFRmut+ NSCLC. Further investigation for the underlying mechanism is warranted. Clin Cancer Res; 21(22); 5173–80. ©2015 AACR.

Risk factors for pregnancy-associated breast cancer: a report from the Nigerian Breast Cancer Study

PURPOSE Little is known about risk factors for pregnancy-associated breast cancer (PABC), diagnosed during pregnancy or postpartum. METHODS We enrolled 1715 premenopausal women from the Nigerian Breast Cancer Study from 1998 to 2011. Based on recency of last pregnancy from diagnosis, breast cancer cases were categorized as (1) PABC diagnosed 2 years or longer postpartum, (2) PABC diagnosed 3 to 5 years postpartum, or (3) non-PABC diagnosed more than 5 years postpartum. Controls were matched to cases on recency of last pregnancy. Multiple logistic regressions were performed comparing cases and controls within each group. RESULTS Of the 718 cases, 152 (21.2%) had PABC 2 or more years postpartum, and 145 (20.2%) 3 to 5 years postpartum. Although not statistically significant, women with higher parity tend to have an elevated risk of PABC but reduced risk of non-PABC (p for heterogeneity = 0.097). Family history of breast cancer might be a strong predictor particularly for PABC 2 or more years postpartum (odds ratio, 3.28; 95% confidence interval, 1.05-10.3). Compared with non-PABC cases, PABC 2 or more years postpartum cases were more likely to carry BRCA1/2 mutations (P = .03). CONCLUSIONS Parity may have different roles in the development of PABC versus other premenopausal breast cancer in Nigerian women. Prospective mothers with multiple births and a family history of breast cancer may have an elevated risk of breast cancer during their immediate postpartum period.

Microsatellites in the Estrogen Receptor (ESR1, ESR2) and Androgen Receptor (AR) Genes and Breast Cancer Risk in African American and Nigerian Women

Genetic variants in hormone receptor genes may be crucial predisposing factors for breast cancer, and microsatellites in the estrogen receptor (ESR1, ESR2) and androgen receptor (AR) genes have been suggested to play a role. We studied 258 African-American (AA) women with breast cancer and 259 hospital-based controls, as well as 349 Nigerian (NG) female breast cancer patients and 296 community controls. Three microsatellites, ESR1_TA, ESR2_CA and AR_CAG, in the ESR1, ESR2 and AR genes, respectively, were genotyped. Their repeat lengths were then analyzed as continuous and dichotomous variables. Analyses of continuous variables showed no association with breast cancer risk in either AA or NG at ESR1_TA; AA cases had shorter repeats in the long allele of ESR2_CA than AA controls (Mann-Whitney P  = 0.036; logistic regression P  = 0.04, OR  = 0.91, 95% CI 0.83–1.00), whereas NG patients had longer repeats in the short allele than NG controls (Mann-Whitney P  = 0.0018; logistic regression P  = 0.04, OR  = 1.06, 95% CI 1.00–1.11); and AA cases carried longer repeats in the short allele of AR_CAG than AA controls (Mann-Whitney P  = 0.038; logistic regression P  = 0.03, OR  = 1.08, 95% CI 1.01–1.15). When allele sizes were categorized as dichotomous variables, we discovered that women with two long alleles of ESR2_CA had increased risk of breast cancer (OR  = 1.38, 95% CI 1.10–1.74; P  = 0.006). This is the first study to investigate these three microsatellites in hormonal receptor genes in relation to breast cancer risk in an indigenous African population. After adjusting for multiple-testing, our findings suggest that ESR2_CA is associated with breast cancer risk in Nigerian women, whereas ESR1_TA and AR_CAG seem to have no association with the disease among African American or Nigerian women.